Histologic and Molecular Characterization of Lung Cancer With Tissue Obtained by Electromagnetic Navigation Bronchoscopy

Ha, Duc MD*; Choi, Humberto MD; Almeida, Francisco A. MD, MS; Arrossi, Andrea MD; Brainard, Jennifer MD; Cicenia, Joseph MD; Farver, Carol MD; Gildea, Thomas MD, MS; Machuzak, Michael S. MD; Mazzone, Peter MD, MPH

Journal of Bronchology & Interventional Pulmonology:
doi: 10.1097/LBR.0b013e31828197e9
Original Investigations

Background: Electromagnetic navigation bronchoscopy (ENB) is a catheter-based adjunct to standard bronchoscopic techniques for the sampling of lung lesions. We sought to evaluate the adequacy of ENB-obtained samples for histologic subtyping of lung cancer, epidermal growth factor receptor (EGFR) mutations, and echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) translocations.

Methods: We retrospectively analyzed consecutive patients who underwent ENB for the diagnosis of lung lesions between 2008 and 2011. In those proven to be a primary lung cancer by ENB, tissue adequacy for histologic subtyping was recorded. Accuracy was determined by comparison with resected specimens when available. Tissue adequacy for EGFR mutation and/or EML4-ALK analyses was also reviewed.

Results: Sixty-five ENB cases resulted in a diagnosis of lung cancer. Tissues obtained were adequate for histologic subtyping in all 65 cases. Forty-three (66.2%) were diagnosed with adenocarcinoma, 19 (29.2%) with squamous cell carcinoma, 3 (4.6%) with small cell carcinoma. In 51 cases (78.5%), subtyping was performed by morphology alone, whereas 11 (21.5%) required immunohistochemical staining. Sixteen of 65 tumors underwent surgical resection. Concordance of histologic subtyping between ENB and surgical specimens was 87.5% (14 tumors). ENB-obtained samples from 15 patients with adenocarcinoma were sent for EGFR mutation analysis, of which 14 (93.3%) were adequate. Samples from 2 patients were evaluated for EML4-ALK gene rearrangements, both of which were adequate for analysis.

Conclusions: ENB is effective at obtaining tissue samples adequate for histologic subtyping, EGFR mutation, and EML4-ALK translocation analysis.

Author Information

*Department of Internal Medicine

Respiratory Institute

Pathology and Laboratory Medicine Institute, Cleveland Clinic, Cleveland, OH

Disclosure: There is no conflict of interest or other disclosures.

Reprints: Duc Ha, MD, Department of Internal Medicine, Cleveland Clinic; 9500 Euclid Avenue, NA10, Cleveland, OH 44195 (e-mail: had@ccf.org).

Received August 25, 2012

Accepted December 4, 2012

Article Outline

Histologic subtyping plays an important role in deciding the course of treatment for patients diagnosed with lung cancer.1 Previous studies have shown that patients respond differently to chemotherapy and targeted therapy depending on the histologic subtypes of their lung cancer.2,3 For instance, patients with stage 4 nonsquamous non-small cell lung cancer (NSCLC) have improved survival and less toxicity when treated with pemetrexed combined with a platinum agent compared with other chemotherapy agents.3,4 Thus, histologic subtyping of NSCLC is now the standard of care for the treatment of lung cancer.5–7 The International Association for the Study of Lung Cancer, American Thoracic Society, and European Respiratory Society recommends that for small biopsy/cytology samples, NSCLCs should be classified into their respective histologic subtypes according to morphologic patterns. When a clear morphologic pattern is not present, immunohistochemical (IHC) staining is suggested to assist with the subtyping.8

In addition to histologic subtyping, treatment of metastatic lung cancer is also guided by the detection of genetic alterations. In patients with metastatic NSCLC who harbor an epidermal growth factor receptor (EGFR) gene mutation, targeted therapy with EGFR tyrosine kinase inhibitors (erlotinib or gefitinib) are recommended as first-line treatment because of greater response rates and longer progression-free survival compared with standard chemotherapeutic regimens.9–12

In addition, in patients with echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) rearrangements, treatment with the tyrosine kinase inhibitor crizotinib leads to a high response rate and improved survival.13,14

For these reasons, the ability to obtain tissue adequate for histologic subtyping and molecular analyses is a key component of the evaluation of the lung cancer patient. Electromagnetic navigation is a guidance technology developed to assist with the localization of peripheral lung lesions during bronchoscopy.15,16 Previous studies have reported the diagnostic yield of electromagnetic navigation bronchoscopy (ENB) to be 63% to 74% for peripheral lung lesions.15–18 The pneumothorax rate for ENB has been reported to be between 1.2% and 6%, significantly lower than the pooled 15% rate for transthoracic needle aspiration (TTNA).16,19–21 Here, we aim to assess the accuracy and adequacy of diagnostic samples obtained by ENB for histologic subtyping and molecular characterization of lung cancer.

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We conducted a retrospective analysis of consecutive patients who, between August 2008 and December 2011, underwent outpatient ENB at the Cleveland Clinic Main Campus that resulted in the diagnosis of lung cancer. The study was approved by the Cleveland Clinic Institutional Review Board.

Study data including patient and lung lesion characteristics were collected and managed using REDCap electronic data capture tools hosted at Cleveland Clinic.22 Tumor size, shape, and location were extracted from radiology reports, and when absent, determined by reviewing original computed tomography images. Clinical staging according to the 7th edition of the TNM staging system was recorded, and when not documented, determined by reviewing available imaging reports.23

Cytopathology reports were reviewed for histologic subtyping of lung cancer, and bronchoscopy reports for the diagnostic sampling methods.24 Histologic subtyping was documented as “adenocarcinoma,” “NSCLC, favor adenocarcinoma,” “squamous cell carcinoma,” “NSCLC, favor squamous cell,” “non–small cell carcinoma, not otherwise specified (NOS),” and “small cell carcinoma.” In cases in which the subtype was not determined by morphology alone, IHC stains for TTF-1, p63, and cytokeratin 5/6 were documented when performed. Histologic subtyping of samples obtained by ENB and surgical resection were recorded separately and compared for concordance.

The adequacy of samples for EGFR mutation (real-time-PCR) and EML4-ALK rearrangement (fluorescence in situ hybridization) analyses was determined by the presence of a result or statement that the sample was inadequate in the test report. EGFR testing was carried out at an outside facility (Clarient, Aliso Vieja, CA).

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Statistical Analysis

Descriptive statistics for all continuous variables were summarized as means, median, and ranges. Statistical analyses were performed using the commercially available software SPSS, version 17 (SPSS Inc., Chicago, IL).

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Sixty-five patients with lung cancer diagnosed by ENB were identified. Their mean age was 71 years (44 to 89 y). Thirty-four patients (52.3%) were females. Twenty-one patients (32.3%) were current smokers, and 41 (63.1%) were former smokers. The median long-axis diameter of the nodules/masses sampled was 24 mm (12 to 57 mm). Fifty-nine (90.8%) were either irregular, lobular, or spiculated. Thirty-seven (56.9%) had clinical stage I or II lung cancer at the time of bronchoscopy (Table 1).

Methods of tissue sampling performed during ENB included brushing, transbronchial biopsy (TBBx) (SuperTrax biopsy forceps, tip diameter 1.7 mm, length 110 cm), 21-G SuperTrax needle aspiration, 22-G Wang needle aspiration, and bronchoalveolar washing. In cases in which only 1 sampling method was diagnostic, TBBx was the most common method (n=8) (Table 2).

A histologic subtype was identified in all 65 cases (100%). Three cases (4.6%) were diagnosed as small cell carcinoma and 62 cases (95%) as NSCLC, of which 51 (78.5%) could be subclassified by morphology only—35 (81.4%) as adenocarcinomas and 16 (84.2%) as squamous cell carcinomas (Table 3). Twenty-two IHC stains were used in 11 (21.5%) cases, of which 8 (18.6%) were NSCLC, favored adenocarcinomas and 3 (15.8%) were NSCLC, favored squamous cell carcinomas. All 22 samples (100%) were adequate for IHC analysis (Table 4). Histologic subtype was concordant between sampling methods when <1 sampling method yielded a diagnosis. For the entire cohort, IHC staining was performed on samples obtained by TBBx (n=12), 21-G SuperTrax needle aspiration (n=1), and 22-G Wang needle aspiration (n=2).

Of the 43 cases of adenocarcinoma classified by ENB, 15 were sent for EGFR mutation analysis. Fourteen (93.3%) were considered adequate for analysis. Two cases were sent for EML4-ALK gene translocation testing, both of which were adequate (Table 4). EGFR mutation testing and EML4-ALK analyses were performed on TBBx samples in all cases except for one in which EGFR mutation analysis was performed on a 22-G Wang needle aspiration sample. The 1 sample inadequate for EGFR mutation analysis was sent from a TBBx sample.

Of the 37 patients with stage I or II lung cancer, 21 were deemed poor surgical candidates because of comorbidities and did not undergo surgical resection of the primary tumor. Sixteen patients underwent surgical resection. Of the 14 cases of ENB classified adenocarcinoma that underwent surgical resection, 13 were confirmed as adenocarcinoma (92.9% concordance). One ENB classified as “NSCLC, favor adenocarcinoma” was found to be a large cell carcinoma after resection and IHC stains performed on the surgical sample. Of the 2 cases of ENB-classified squamous cell carcinomas that underwent surgical resection, one was found to be an adenosquamous carcinoma after resection. In total, there was an 87.5% concordance rate between histologic subtypes obtained by ENB compared with those obtained by surgical resection (Table 3). In the 2 cases that were misclassified by ENB, IHC staining was not performed on the ENB-obtained tissue samples. IHC staining was performed on the surgical specimen for the adenosquamous carcinoma. It was reported as predominantly squamous cell carcinoma based on morphology. In the 1 ENB subtyped adenocarcinoma that was a large cell carcinoma on surgical resection, brushings alone were used to obtain tissue for diagnosis. Tissue from the adenosquamous carcinoma that was subtyped as squamous cell carcinoma on ENB was obtained by brushing, 21-G SuperTrax needle aspiration, and TBBx.

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In this study, we sought to determine the adequacy of tissue samples obtained by ENB for histologic subtyping, EGFR mutation, and EML4-ALK translocation analyses. In 65 cases of lung cancer diagnosed by ENB, the samples obtained were adequate for histologic subtyping of lung cancer in all cases. In addition, the histologic subtyping was accurate, with an 87.5% (14/16) concordance rate with surgical specimens. In samples where morphology alone was not able to provide the subtype of lung cancer, IHC staining was able to differentiate between the subtypes. All tissue samples submitted for IHC staining were adequate for the necessary analyses. In addition, 94.1% (16/17) of the samples tested were adequate for EGFR mutation or EML4-ALK translocation analyses.

Previous reports have indicated small biopsy/cytology samples obtained by TTNA, and endobronchial ultrasound-guided transbronchial needle aspiration are adequate for histologic and molecular characterization of lung cancer.25–29 To the best of our knowledge, 2 previous reports have described the adequacy of samples obtained by standard bronchoscopy for IHC staining to subtype lung cancer. In both the studies, specimens were obtained either by bronchial washing or brushing.30,31 ENB is an alternative to TTNA in the evaluation of peripheral, smaller lung lesions, and has been previously described to be effective and safe.15–18 This is the first report describing the effectiveness of ENB in obtaining tissue adequate for histologic subtyping, EGFR mutation, and EML4-ALK translocation analysis. The histologic subtypes of lung cancer from tissues obtained by ENB were accurate when compared with those obtained by surgical resection. The squamous cell carcinoma classified by ENB underwent IHC staining on the surgical sample to confirm the adenocarcinoma component. This suggests that small samples obtained by various bronchoscopic techniques, including ENB, may be inadequate for subtyping mixed tumors because of selective sampling of the tumor. Tissue samples obtained by ENB are not expected to be different from those obtained by traditional bronchoscopy in the evaluation of peripheral lesions, as the methods of sampling tend to be the same.

Histologic subtyping and molecular characterization of late-stage lung cancer is currently required to provide standard of care treatment. Standard chemotherapy regimens are now based on histology, and histology guides the need for EGFR mutation analysis and EML4-ALK testing.1,8 Agents targeting pathways affected by EGFR activating mutations and the EML4-ALK translocation are considered standard of care therapies when alterations are confirmed. A full 45% of patients in our series had stage III or IV disease at the time of the diagnostic ENB.

Many patients with early-stage lung cancer are not well enough to tolerate resection and instead are treated with ablative therapies. Early recurrence after curative intent treatment may occur in locations that are difficult to biopsy. At times, ENB may be the easiest and safest means of obtaining tissue in this group. Although chemotherapy drugs may have limited role in those with early-stage lung cancer treatable with radiation therapy, mutational analysis clearly has value in those with recurrent, metastatic disease.

The importance of histologic subtyping and molecular characterization of early-stage lung cancer is likely to grow over time. Advances in molecular biomarker development may lead to an improved understanding of the need for adjuvant chemotherapy or chemoprevention. In addition, biomarkers capable of prognosticating outcomes and predicting responses to therapy are under development. The results obtained in this study suggest that ENB-obtained samples will be adequate for both histologic subtyping and molecular characterization as future tests are developed.

Our study is limited by its retrospective nature. Not all samples obtained by ENB were sent for IHC staining, analysis of EGFR mutations and EML4-ALK translocations, and the outcomes of interest were on the basis of the review of cytopathology reports. We were not able to assess the concordance of molecular analyses with those performed on surgical specimens. The increased use of endobronchial ultrasound-guided transbronchial needle aspiration for advanced stage disease with mediastinal adenopathy was not reviewed in this study as we set out only to understand the value in peripheral lung sampling.

We conclude that ENB is effective at obtaining tissue samples adequate for histologic subtyping of lung cancer, as well as EGFR mutation and EML4-ALK translocation analysis.

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1. Bareschino MA, Schettino C, Rossi A, et al. Treatment of advanced non small cell lung cancer. J Thorac Dis. 2011;3:122–133
2. Sandler A, Gray R, Perry MC, et al. Paclitaxel-carboplatin alone or with bevacizumab for non-small-cell lung cancer. N Engl J Med. 2006;355:2542–2550
3. Scagliotti GV, Parikh P, von Pawel J, et al. Phase III study comparing cisplatin plus gemcitabine with cisplatin plus pemetrexed in chemotherapy-naive patients with advanced-stage non-small-cell lung cancer. J Clin Oncol. 2008;26:3543–3551
4. Scagliotti G, Brodowicz T, Shepherd FA, et al. Treatment-by-histology interaction analyses in three phase III trials show superiority of pemetrexed in nonsquamous non-small cell lung cancer. J Thorac Oncol. 2011;6:64–70
5. Bulman W, Saqi A, Powell CA. Acquisition and processing of endobronchial ultrasound-guided transbronchial needle aspiration specimens in the era of targeted lung cancer chemotherapy. Am J Respir Crit Care Med. 2012;185:606–611
6. Cooper WA, O’toole S, Boyer M, et al. What’s new in non-small cell lung cancer for pathologists: the importance of accurate subtyping, EGFR mutations and ALK rearrangements. Pathology. 2011;43:103–115
7. Mukhopadhyay S. Utility of small biopsies for diagnosis of lung nodules: doing more with less. Mod Pathol. 2012;25(suppl 1):S43–S57
8. Travis WD, Brambilla E, Noguchi M, et al. International association for the study of lung cancer/American thoracic society/European respiratory society international multidisciplinary classification of lung adenocarcinoma. J Thorac Oncol. 2011;6:244–285
9. Mitsudomi T, Morita S, Yatabe Y, et al. Gefitinib versus cisplatin plus docetaxel in patients with non-small-cell lung cancer harbouring mutations of the epidermal growth factor receptor (WJTOG3405): an open label, randomised phase 3 trial. Lancet Oncol. 2010;11:121–128
10. Mok TS, Wu YL, Thongprasert S, et al. Gefitinib or carboplatin-paclitaxel in pulmonary adenocarcinoma. N Engl J Med. 2009;361:947–957
11. Maemondo M, Inoue A, Kobayashi K, et al. Gefitinib or chemotherapy for non-small-cell lung cancer with mutated EGFR. N Engl J Med. 2010;362:2380–2388
12. Cataldo VD, Gibbons DL, Perez-Soler R, et al. Treatment of non-small-cell lung cancer with erlotinib or gefitinib. N Engl J Med. 2011;364:947–955
13. Kwak EL, Bang YJ, Camidge DR, et al. Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer. N Engl J Med. 2010;363:1693–1703
14. Shaw AT, Yeap BY, Solomon BJ, et al. Effect of crizotinib on overall survival in patients with advanced non-small-cell lung cancer harbouring ALK gene rearrangement: a retrospective analysis. Lancet Oncol. 2011;12:1004–1012
15. Gildea TR, Mazzone PJ, Karnak D, et al. Electromagnetic navigation diagnostic bronchoscopy: a prospective study. Am J Respir Crit Care Med. 2006;174:982–989
16. Schwarz Y. Electromagnetic navigation. Clin Chest Med. 2010;31:65–73
17. Makris D, Scherpereel A, Leroy S, et al. Electromagnetic navigation diagnostic bronchoscopy for small peripheral lung lesions. Eur Respir J. 2007;29:1187–1192
18. Eberhardt R, Anantham D, Herth F, et al. Electromagnetic navigation diagnostic bronchoscopy in peripheral lung lesions. Chest. 2007;131:1800–1805
19. Eberhardt R, Anantham D, Ernst A, et al. Multimodality bronchoscopic diagnosis of peripheral lung lesions: a randomized controlled trial. Am J Respir Crit Care Med. 2007;176:36–41
20. Wang Memoli JS, Nietert PJ, Silvestri GA. Meta-analysis of guided bronchoscopy for the evaluation of the pulmonary nodule. Chest. 2012;142:385–393
21. Wiener RS, Schwartz LM, Woloshin S, et al. Population-based risk for complications after transthoracic needle lung biopsy of a pulmonary nodule: an analysis of discharge records. Ann Intern Med. 2011;155:137–144
22. Harris PA, Taylor R, Thielke R, et al. Research electronic data capture (REDCap)—a metadata-driven methodology and workflow process for providing translational research informatics support. J Biomed Inform. 2009;42:377–381
23. Edge SB, Byrd DR, Compton CC, et al, eds. Lung. American Joint Committee on Cancer Staging Manual. 7th ed. New York: Springer; 2010. p. 253
24. Travis WD, Brambilla E, Muller-Hermlink HK. eds. Pathology and genetics of tumours of the lung, pleura, thymus and heart. World Health Organization Classification of Tumours. 2004 Lyon IARC Press:p. 9–122
25. Cheung YC, Chang JW, Hsieh JJ, et al. Adequacy and complications of computed tomography-guided core needle biopsy on non-small cell lung cancers for epidermal growth factor receptor mutations demonstration: 18-gauge or 20-gauge biopsy needle. Lung Cancer. 2010;67:166–169
26. Garcia-Olive I, Monso E, Andreo F, et al. Endobronchial ultrasound-guided transbronchial needle aspiration for identifying EGFR mutations. Eur Respir J. 2010;35:391–395
27. Nakajima T, Yasufuku K, Suzuki M, et al. Assessment of epidermal growth factor receptor mutation by endobronchial ultrasound-guided transbronchial needle aspiration. Chest. 2007;132:597–602
28. Navani N, Brown JM, Nankivell M, et al. Suitability of endobronchial ultrasound-guided transbronchial needle aspiration specimens for subtyping and genotyping of non-small cell lung cancer: a multicenter study of 774 patients. Am J Respir Crit Care Med. 2012;185:1316–1322
29. Solomon SB, Zakowski MF, Pao W, et al. Core needle lung biopsy specimens: adequacy for EGFR and KRAS mutational analysis. AJR Am J Roentgenol. 2010;194:266–269
30. Yang Y, Pan QJ, Teng MF, et al. Application of protein markers in combination with ThinPrep bronchial brush cytology in classification of lung cancer subtypes. Zhonghua Zhong Liu Za Zhi. 2008;30:616–619
31. Collins GR, Thomas J, Joshi N, et al. The diagnostic value of cell block as an adjunct to liquid-based cytology of bronchial washing specimens in the diagnosis and subclassification of pulmonary neoplasms. Cancer Cytopathol. 2012;120:134–141

electromagnetic navigation bronchoscopy; lung cancer; histology; immunohistochemistry; epidermal growth factor receptor (EGFR) mutation; echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) translocation

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